When organisms learn, their memory can last either for a short or long- period of time. Studies show that short-term memory (lasting minutes to hours) involves covalent modifications of pre-existing proteins. In contrast, long-term memory (lasting hours to days) appears to involve new protein synthesis and gene expression. Inhibition of translation and transcription during the acquisition phase blocks the formation of long- term memory without any effect on short-term memory. Moreover, long-term memory is often accompanied and perhaps mediated by morphological changes in neurons. The biochemical events leading to long-term memory and the associated morphological changes are virtually unknown. While a few genes and proteins have been shown to be altered in their expression following long-term training, it has been difficult to investigate causal relationships between long-term memory formation and the functions of these identified genes and proteins. The purpose of this project is to identify and characterize transcription factors and immediate-early genes activated by long-term training and to investigate their functional role in memory formation and the associated morphological changes. We will carry out these experiments in the marine mollusc Aplysia californica. Because of its simple and accessible nervous system, Aplysia has been one of the major systems used in memory research. A critical locus for the storage of memory for long-term sensitization of the gill- and siphon- withdrawal reflex is the connection between the sensory neurons and the motor neurons which can be reconstituted in dissociated cell cultures. Previous studies in intact animals and in sensory-motor neuron cultures indicate that cAMP mediated gene induction may be necessary for long-term memory formation as well as the associated morphological changes in Aplysia. The project has four specific aims. Aim 1 is to test the hypothesis that the transcription factor CREB (cAMP responsive element binding protein) is part of the biochemical pathway by which long-term memory is induced in Aplysia. Aim 2 is to test the hypothesis that immediate-early genes are involved in the induction of long-term memory. Aim 3 is to test the hypothesis that morphological changes seen following long-term training are caused by activation of CREB protein. Aim 4 is to isolate the cDNA clone encoding Aplysia CREB protein and to test the hypothesis that microinjection of the Aplysia protein into sensory neurons can induce long-term facilitation and the associated morphological changes. A combination of biochemical, electrophysiological and behavioral experiments in intact animals, in semi-intact ganglia and in sensory-motor neuron cultures will be to test the above hypotheses. Understanding the biochemical pathway(s) involved in the formation of memory will provide insights into mechanisms responsible for establishment of long-lasting plasticities in the brain. This information also would be of great value in understanding the consequences of some morphological diseases associated with memory deficits as well as injury to the brain.